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ObjectiveTo investigate the protective effect of the extract of Liuwei Dihuangwan （LW） on mitochondrial damage in the Alzheimer's disease （AD） model of Caenorhabditis elegans （C. elegans）. MethodC. elegans transfected with human β-amyloid protein （Aβ） 1-42 gene was used as an AD model. The rats were divided into blank group， model group， metformin group （50 mmol·L-1）， and low， medium， and high dose （1.04， 2.08， 4.16 g·kg-1） LW groups. Behavioral methods were used to observe the sensitivity of 5-hydroxytryptamine （5-HT） in nematodes. Western blot was used to detect the expression of Aβ in nematodes. Total ATP content in nematodes was detected by the adenine nucleoside triphosphate （ATP） kit， and mitochondrial membrane potential was detected by the JC-1 method. In addition， the mRNA expression of Aβ expression gene （Amy-1）， superoxide dismutase-1 （SOD-1）， mitochondrial transcription factor A homologous gene-5 （HMG-5）， mitochondrial power-associated protein 1 （DRP1）， and mitochondrial mitoprotein 1 （FIS1） was detected by real-time fluorescence quantitative polymerase chain reaction （RT-PCR）. ResultThe extract of LW could reduce the hypersensitivity of the AD model of nematodes to exogenous 5-HT （P<0.05） and delay the AD-like pathological characteristics of hypersensitivity to exogenous 5-HT caused by toxicity from overexpression of Aβ in neurons of the AD model of nematodes. Compared with the blank group， in the model group， the mRNA expression of Aβ protein and Amy-1 increased （P<0.01）， and the mRNA expression of SOD-1 and HMG-5 decreased （P<0.01）. The mRNA expression of DRP1 and FIS1 increased （P<0.01）， and the level of mitochondrial membrane potential decreased （P<0.05）. The content of ATP decreased （P<0.01）. Compared with the model group， in the positive medicine group and medium and high dose LW groups， the mRNA expression of Aβ protein and Amy-1 decreased （P<0.05，P<0.01）， and the mRNA expression of SOD-1 and HMG-5 increased （P<0.01）. The mRNA expression of DRP1 decreased （P<0.05，P<0.01）， and that of FIS1 decreased （P<0.01）. The level of mitochondrial membrane potential increased （P<0.01）， and the content of ATP increased （P<0.05，P<0.01）. ConclusionThe extract of LW may enhance the antioxidant ability of mitochondria， protect mitochondrial DNA， reduce the fragmentation of mitochondrial division， repair the damaged mitochondria， adjust the mitochondrial membrane potential， restore the level of neuronal ATP， and reduce the neuronal damage caused by Aβ deposition.
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ObjectiveTo study the efficacy of Aclasta combined with Liuwei Dihuangwan on osteoporosis and the effect on quality of life. MethodA total of 126 patients with osteoporosis who were treated in the Affiliated Hospital of Shandong University of Traditional Chinese Medicine from September 2019 to September 2020 were classified into the observation group and the control group with the randomized double-blind method. The observation group consisted of 60 patients （26 males and 34 females） with the age of 59-85 years old ［mean： （72.0 ± 6.5） years old］. The control group was composed of 66 patients （31 males and 35 females）， with the age of 62-82 years old ［mean： （73.0±8.2） years old］. The control group was treated with Aclasta， and the observation group Aclasta combined with Liuwei Dihuangwan. After treatment， the effective rate of each group was calculated. Bone mineral density （BMD） was measured in both groups before and after treatment， and serological parameters calcium （Ca）， total 25 （OH） vitamin D3 （VITD-T）， osteocalcin （OC）， serum alkaline phosphatase （ALP）， parathyroid hormone （PTH）， β-collagen special sequence （β-CTX）， and total procollagen 1 N-terminal propeptide （T-P1NP） were also measured. Visual Analogue Scale （VAS） score， Japanese Orthopaedic Association （JOA） score， and Oswestry Disability Index （ODI） score were evaluated. On this basis， the effect was compared between the two groups. ResultThe indexes were insignificantly different between the two groups before treatment. After 6 months of treatment， the two groups showed decrease in VAS score and ODI score （P<0.01）， increase in JOA score （P<0.01）， BMD of lumbar spine and hip joint， elevation of Ca， VITD-T， OC， ALP， and PTH （P<0.05， P<0.01）， and decrease of β-CTX （P<0.01） as compared with before treatment. The level of T-P1NP dropped in the observation group after treatment （P<0.01）.After treatment， the total effective rate of the observation group was 88.3% （53/60）， as compared with the 74.2% （49/66） in the control group （χ2=4.047， P<0.05）. Moreover， after treatment， the observation group demonstrated higher levels of BMD， Ca， VITD-T， OC， and PTH （P<0.05）， lower levels of T-P1NP （P<0.05）， lower VAS score （P<0.01）， and higher JOA score （P<0.05） than the control group， but the ODI score was insignificantly different from that in the control group. ConclusionAclasta combined with Liuwei Dihuangwan is effective on osteoporosis， without increasing the incidence of adverse reactions. In addition， the combination can alleviate pain and improve the quality of life of osteoporosis patients.
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ObjectiveTo evaluate the efficacy of Zishui Bugan decoction on perimenopausal insomnia of liver-kidney deficiency type and the safety. MethodA randomized， double-blind， placebo-controlled trial was designed. To be specific， 72 patients of perimenopausal insomnia with Liver-kidney deficiency were randomized into the treatment group and the control group at the ratio of 1∶1 and they were respectively treated with Zishui Bugan decoction and placebo for 3 weeks. The Pittsburgh sleep quality index （PSQI）， modified Kupperman index， traditional Chinese medicine （TCM） syndrome score， self-rating anxiety scale （SAS） score， and self-rating depression scale （SDS） score， were compared before and after treatment to determine the clinical efficacy， with adverse effects recorded. ResultThe total effective rate for insomnia was 85.3%（29/34） in treatment group and 17.6%（6/34） in control group（Z=-5.582，P<0.01）. After treatment， PSQI score， modified Kupperman index， TCM syndrome score， and SAS score were improved in both groups （P<0.05， P<0.01）， particularly the treatment group which showed significant difference from the control group （P<0.05，P<0.01）. The safety indicators were insignificantly different between two groups before and after treatment. No related adverse effects were reported in both groups during the treatment. ConclusionZishui Bugan decoction can improve the sleep quality and alleviate the menopausal symptoms， such as depression and anxiety， which shows ideal efficacy and safety for the perimenopausal insomnia with liver-kidney deficiency type.
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ObjectiveTo investigate the protective effect of Liuwei Dihuangwan on neurovascular injury in SAMP8 mice. MethodThe Alzheimer's disease （AD） model with insufficiency of kidney essence was induced in 75 SAMP8 mice aging 6 months. The model mice were divided into model group， positive control group （donepezil hydrochloride， 0.747 mg·kg-1·d-1）， and high-， medium-， and low-dose Liuwei Dihuangwan groups （2.700， 1.350， 0.675 g·kg-1·d-1）， with 15 mice in each group. Fifteen SAMR1 mice were assigned to a normal control group. All mice were administered continuously for 2 months. The spatial memory of mice was tested by the Morris water maze. Hematoxylin-eosin （HE） staining was used to observe the pathological changes in the hippocampus and cortex of brain tissues. The immunohistochemical method （IHC） was used to detect the deposition of amyloid β-protein （Aβ） and the expression of von Willebrand factor （vWF） and CD34 in the hippocampus and cortex of brain tissues. Electron microscopy was used to observe the ultrastructural changes in cerebral microvessels. Western blot was used to detect the protein expression levels of the receptor of advanced glycation endproduct （RAGE）， low-density lipoprotein receptor-related protein 1 （LRP1）， vascular endothelial growth factor A （VEGF-A）， and P-selection in the hippocampus and cortex of brain tissues. ResultCompared with the normal control group， the model group showed prolonged escape latency and swimming distance （P<0.01）， increased number of glial cells， decreased number of nerve cells， blurred tight junctions or enlarged gap of the brain microvascular endothelial cells， severely injured membrane structure， swollen mitochondria of endothelial cells， ruptured membrane， massive dissolution in cristae， increased protein expression of Aβ and vWF in the hippocampus and cortex （P<0.01）， reduced protein expression of CD34 （P<0.05）， elevated protein expression of RAGE and P-selection in the cortex （P<0.01）， and decreased protein expression level of LRP1 and VEGF-A （P<0.01）. Compared with the model group， the Liuwei Dihuangwan groups showed shortened escape latency and swimming distance （P<0.05）， reduced number of glial cells in the cortex and hippocampus， increased number of microvessels in the cortex， clear double-layer membrane structure in tight junctions between the microvascular endothelial cells， increased number of mitochondria with intact membrane and recovered mitochondrial cristae， decreased protein expression of Aβ， vWF， RAGE， and P-selection in the hippocampus and cortex （P<0.05）， and increased protein expression of CD34， LRP1， and VEGF-A （P<0.05）. ConclusionLiuwei Dihuangwan can regulate Aβ metabolism through the RAGE/LRP1 receptor system and promote cerebral microvascular angiogenesis by inhibiting vWF expression and increasing VEGF-A and CD34， thereby improving cerebral microvascular injury in SAMP8 mice.
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ObjectiveTo investigate the effect of Liuwei Dihuangwan on memory function of senescence-accelerated mouse prone 8 （SAMP8） mice by regulating autophagy through the phosphatidylinositol 3-kinase （PI3K）/protein kinase B （Akt）/forkhead box O3a （FoxO3a） pathway. MethodSix male senescence-accelerated mouse resistant 1 （SAMR1） mice of SPF grade aging 6 months were assigned to a normal group， and 24 male SAMP8 mice of SPF grade aging 6 months were randomly divided into a model group， a donepezil group （0.747 mg·kg-1）， and high- and low-dose Liuwei Dihuangwan groups （2.700 and 1.350 g·kg-1）， with 6 mice in each group. The mice were treated with drugs by gavage for 2 months. Morris water maze was used to detect the learning and memory abilities of mice in each group. Nissl staining was used to observe the neurons in the cortex and hippocampus. The positive expression of microtubule-associated protein 1 light chain 3B （LC3B） in the cortex and hippocampus was detected by immunohistochemistry （IHC）. Western blot was used to detect the protein expression of the mammalian ortholog of yeast ATG6 （Beclin-1）， B cell lymphoma-2 （Bcl-2）， autophagy-related gene 5 （ATG5）， cysteinyl aspartate-specific protease 3 （Caspase-3）， Caspase-9， Akt， p-Akt， FoxO3a， and p-FoxO3a. ResultCompared with the normal group， the model group showed prolonged escape latency （P<0.05，P<0.01）， reduced number of platform crossings and the residence time in the target quadrant （P<0.01）， decreased neurons with reduced volume and dispersed distribution in the cortex and hippocampus， increased positive expression of LC3B （P<0.01）， elevated expression of Beclin-1 and ATG5 in the cortex （P<0.01）， declined Bcl-2 expression （P<0.01）， up-regulated Caspase-3 and Caspase-9 expression （P<0.01）， and decreased expression levels of p-Akt/Akt and p-FoxO3a/FoxO3a （P<0.01）. Compared with the model group， the donepezil group and the Liuwei Dihuangwan groups showed shortened 3 d escape latency （P<0.05，P<0.01）， increased number of platform crossings （P<0.01）， and prolonged residence time in the target quadrant （P<0.01）. In the donepezil group， the number of neurons in the cortex and hippocampus was increased. In the Liuwei Dihuangwan groups， the number of neurons and Nissl bodies increased with denser distribution and lower degree of cell damage. The positive expression of LC3B in the cortex and hippocampus was decreased in the donepezil group and Liuwei Dihuangwan groups （P<0.01）. The expression of Beclin-1 was decreased in the Liuwei Dihuangwan groups （P<0.01）. The expression of ATG5 was decreased in the donepezil group and the low-dose Liuwei Dihuangwan group （P<0.01）. The donepezil group and the Liuwei Dihuangwan groups showed the increased expression level of Bcl-2 in the cortex （P<0.01）， decreased expression level of Caspase-3 （P<0.01）， reduced expression level of Caspase-9 （P<0.05，P<0.01）， and elevated expression levels of p-Akt/Akt and p-FoxO3a/FoxO3a （P<0.01）. ConclusionLiuwei Dihuangwan can effectively improve the learning and memory abilities of the SAMP8 mice and protect neurons. Its mechanism may be related to the regulation of the PI3K/Akt/FoxO3a signaling pathway， down-regulation of the expression of ATG5， Beclin-1， and LC3B， and the inhibition of apoptosis.
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ObjectiveTo study the underlying mechanism of Liuwei Dihuangwan in inhibiting triple-negative breast cancer through mitogen-activated protein kinase kinase kinase 1 （MAPKKK1） and Krüppel-like factor 4 （KLF4）. MethodFour hundreds SPF female Kunming mice aged 11.5 months were palpated once every 3 days. The model mice of spontaneous tumors were randomly divided into a model group （normal saline）， a paclitaxel group （0.025 g·kg-1·d-1， ip， 21 days）， and high-， medium- and low-dose Liuwei Dihuangwan groups （7.2， 3.6， 1.8 g·kg-1·d-1， ig）. Tumor tissues were separated until the moribund stage. The tumor volume and weight were measured， and the tumor inhibition rate and the survival time of the tumor mice were calculated （after 6 months， tumor-free mice were assigned into the normal group）. SPF SD rats were selected to prepare serum samples containing Liuwei Dihuangwan of different concentrations for cell culture， and MAPKKK1 in MDA-MB-231 cells was silenced. The protein expression of MAPKKK1 and KLF4 was detected by immunofluorescence and Western blot. ResultThe in vivo experimental results showed that compared with the conditions of the normal group， the protein expression of MAPKKK1 and KLF4 in tumor tissues of the model group dropped （P<0.01）. Compared with the model group， all medication groups showed reduced tumor volume and weight （P<0.05， P<0.01）， increased tumor inhibition rate， prolonged survival time of tumor mice （P<0.05）， and increased protein expression of MAPKKK1 （P<0.01）. Additionally， the paclitaxel group and the high-dose Liuwei Dihuangwan group exhibited increased protein expression of KLF4 （P<0.01）. The in vitro experiments showed that compared with the conditions of the normal group， the fluorescence intensities of MAPKKK1 and KLF4 in MDA-MB-231 cells in all medication groups were potentiated， and the protein expression of MAPKKK1 in the paclitaxel group and the high- and medium-dose Liuwei Dihuangwan groups， and the protein expression of KLF4 in the paclitaxel group and high-dose Liuwei Dihuangwan group increased （P<0.01）. After silencing of MAPKKK1， compared with the conditions of the negative plasmid group （unsilenced MAPKKK1）， the fluorescence intensities of MAPKKK1 and KLF4 and the protein expression decreased in the RNAi-27 positive plasmid group （silenced MAPKKK1） （P<0.05， P<0.01）. Compared with the RNAi-27 positive plasmid group， all medication groups had enhanced fluorescence intensities of MAPKKK1 and KLF4 and protein expression to different degrees （P<0.05， P<0.01）. ConclusionLiuwei Dihuangwan can inhibit the growth of triple-negative breast cancer， and the underlying molecular mechanism is related to the up-regulation of MAPKKK1 and activation of KLF4 expression.
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Objective:To observe the effects of Fushengong prescription on p38 mitogen-activated protein kinase（p38 MAPK） signal pathway of rats with chronic renal failure（CRF），and to explore its mechanism of reducing inflammatory reaction of renal tissues and delaying the progress of renal interstitial fibrosis. Method:The 55 male Sprague-Dawley rats were randomly divided into normal group，model group， and low，medium and high dose groups of Fushengong prescription，with 11 rats in each group.The normal group was routinely reared， and the other four groups of rats were fed a diet containing 0.5% adenine to produce a model of CRF， which was continuously molded for 21 days.After successful modeling，all rats switched to conventional feed.Normal group and model group were given normal saline 20 mL·kg-1，and each group of Fushengong prescription was given 4，8，16 g·kg-1 of water prescription once a day for 30 days.After the experiment，Masson staining was used to observe the degree of renal interstitial fibrosis.The expression of monocyte chemotactic protein-1（MCP-1） in renal tissues was detected by immunohistochemistry. The expression of phosphorylated p38 mitogen-activated protein kinase（p-p38 MAPK） and transformed growth factor-β1（TGF-β1） in renal tissues were detected by Western blot. Result:Compared with normal group，the renal interstitial collagen deposition increased significantly，the average optical density value of MCP-1 and the expression levels of p-p38 MAPK and TGF-β1 also increased significantly in model group （P<0.05）. Compared with model group，the renal interstitial collagen deposition reduced significantly，the average optical density value of MCP-1 and the protein expression levels of p-p38 MAPK and TGF-β1 also decreased significantly in each dose group of Fushengong prescription（P<0.05）. Conclusion:Fushengong prescription can effectively inhibit the expression of related inflammatory factors in the renal tissue of CRF rats，so as to reduce the inflammatory response in the renal tissue and delay the progress of renal interstitial fibrosis，the mechanism of which may be related to inhibit the activation of p38 MAPK signal transduction pathway.
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Objective::To discuss the clinical efficacy of Liuwei Dihuangwan combined with Danzi Xiaoyaosan on H-type hypertension with syndrome of Yin deficiency and Yang hyperactivity and its effect on vascular endothelial function and inflammatory factors. Method::One hundred and fifty patients were randomly divided into control group (75 cases) and observation group (75 cases) by random number table.Patients in control group got enalapril maleate and folic acid tablets, 1 tablet/time, 1 time/day, and patients with uncontrollable blood pressure were also given nifedipine sustained-release tablets, 20-30 mg/time, 2 times/days.In addition of the therapy of control group, patients in observation group were also given modified Liuwei Dihuangwan combined with Danzi Xiaoyaosan, 1 dose/day.The course of treatment was 12 weeks.Before and after treatment, systolic blood pressure (SBP), diastolic blood pressure (DBP) and up-to-standard blood pressure were detected.And ambulatory blood pressure, standard deviation of 24-hours systolic blood pressure (24 h SSD), standard deviation of 24-hour diastolic blood pressure 24 h DSD), 24 h mean systolic blood pressure (24 h SBP), 24 h mean diastolic blood pressure (24 h DBP) were recorded.And ankle brachial index (ABI) and brachial artery blood flow mediated diamete (FMD) were discussed, syndrome of Yin deficiency and Yang hyperactivity was scored, and levels of iterleukin-6 (IL-6), IL-10, high-sensitivity C-reactive protein (hsCRP) and homocysteine (Hcy) were detected. Result::The levels of SBP, DBP, 24 h SSD, 24 h DSD, 24 h SBP and 24 h DBP were all lower than those in control group (P<0.01). The achievement rate of accidental blood pressure in observation group was 81.19%, which was higher than 66.98% in control group (χ2=29.81, P<0.01). And levels of ABI, FMD and IL-10 were higher than those in control group (P<0.01), while the score of syndrome of Yin Deficiency and Yang Hyperactivity and the levels of Hcy, leptin, IL-6 and hs-CRP were lower than those in control group (P<0.01). Conclusion::In addition to the therapy of antihypertensive and folic acid, Liuwei Dihuangwan combined with Danzi Xiaoyaosan can be given to control the level of blood pressure and Hcy, relieve the variability of blood pressure, alleviate clinical symptoms, raise the rate of achievement rate of blood pressure, improve the function of vascular endothelium, and regulate inflammatory factors.
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Objective: Taking single Liuwei Dihuangwan(LDW) as the research object,the moisture content change and volumetric shrinkage characteristics in its drying process were investigated,which provided the theoretical basis for improving the drying efficiency of the pills and reducing the quality problems of the pattern pills,crusts and crack pills. Method: The drying characteristics of LDW at drying temperature of 50,75,100,125℃ were studied by constant temperature hot air drying and vacuum drying.Based on sphere model of Fick's second law and Arrhenius equation,the effective diffusion coefficient and activation energy of the drying process were obtained.The volumetric shrinkage characteristics of the pills during the drying process were studied by the projected area method.The Weibull function was used to fit the drying dynamics curve of single LDW. Result: Hot air drying and vacuum drying of LDW both belonged to the decreasing drying processes,and the time required to achieve the same moisture content in vacuum drying was shorter than that in hot air drying.The moisture ratio in the drying process of single LDW obeyed the Weibull function distribution(R2=0.994 5-0.999 7),the scale parameter(α) decreased with the increase of temperature,and drying temperature had significant influence on the shape parameter (β).The effective diffusion coefficients of hot air drying and vacuum drying were 2.626×10-3-7.823×10-2,3.782×10-3-9.042×10-2 m2·s-1, their activation energy were 47.18,42.69 kJ·mol-1,respectively.The volume ratio of hot air drying and vacuum drying of LDW ranged from 0.638 to 0.741 and 0.607 to 0.689,respectively. Conclusion: Weibull function can be adopted to predict the drying and dehydration law of LDW.Under the condition of low temperature drying,slow-down of shrinkage rate of the pills is helpful to prevent the formation of splitting pills,this study provides theoretical and technical basis for dying of LDW.
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Objective: To investigate the protective mechanism of Liuwei Dihuangwan on diabetes mellitus with liver injury in terms of inflammation. Method: The 18 db/db mice were selected from animal model of spontaneous type 2 diabetes, mice were randomly divided into model group, Liuwei Dihuangwan group(9.75 g·kg-1·d-1,once a day), resveratrol group(42.6 mg·kg-1·d-1,once a day)based on fasting blood-glucose (FBG). The 12 litter wild db/m mice were randomly divided into normal group, Liuwei Dihuangwan group(9.75 g·kg-1·d-1,once a day). The normal group and the model group were given the same amount of distilled water by gavage. The mice in each group were treated for 16 weeks. FBG, triglyceride (TG) and alanine aminotransferase (ALT) were examined. Histopathological changes were observed by liver biopsy hematoxylin-eosin (HE) staining. Silent information regulator 6 (SIRT6),nuclear factor-kappaB p65 (NF-κB p65),monocyte chemoattractant protein-1 (MCP-1), vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) in the liver tissue were detected by Western blot. Result: Compared with normal group, FBG, TG and ALT in model group increased significantly(PκB p65,MCP-1,VCAM-1 and ICAM-1 in model group were significantly up-regulated(PPPPκB p65, MCP-1 and VCAM-1 were significantly decreased (PPConclusion: Liuwei Dihuangwan can protect the diabetes mice from liver injury, and its mechanism is related to the improvement of lipid metabolism and the inhibition of inflammatory response.
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Objective: The effect of Liuwei Dihuangwan on blood metabolism during growth and development of rats was investigated by taking the overall metabolic profile and biomarkers of metabolomics as indexes. Method: Ultra performance liquid chromatography-high definition mass spectrometry(UPLC-HDMS) was employed to establish a blood metabolomics study method for characterizing the blood metabolic profile of rats at different time before and after administration. The pattern recognition method was used to integrate and analyze the metabolic profiles, and the differential metabolic markers related to drug action were searched. Based on metabolomics pathway analysis(MetPA) and Kyoto encyclopedia of genes and genomes(KEGG) and other databases, the metabolic pathways related to differential markers were analyzed to study the effect of Liuwei Dihuangwan on blood metabolism during growth and development of rats. Result: Liuwei Dihuangwan significantly affect the blood metabolism profiles during growth and development of rats by regulating 30 blood metabolic markers and 12 related target metabolic pathways, and 8 key metabolic markers were delineated. Conclusion: Liuwei Dihuangwan can significantly regulate the blood metabolic network during the growth and development of rats, thereby affecting the growth and development of rats.
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Objective:To observe the clinical efficacy of addition and subtraction therapy of Liuwei Dihuangwan combined with Huanglian Ejiaotang to primary insomnia in the elderly (liver and kidney Yin deficiency syndrome), and to investigate its regulation effect on neurotransmitter. Method:Randomly 136 patients were divided into control group (68 cases) and observation group (68 cases) by number table. Patients in control group got Estazolam tablets by oral administration before going to bed, 1-2 mg/time, 1 time/day. Patients in observation group got addition and subtraction therapy of Liuwei Dihuangwan combined with Huanglian Ejiao tang, 1 dose/day. Both groups of patients received sleep guidance and cognitive behavior guidance, with treatment course of 8 weeks. During the observation period, physical therapy and acupuncture could not been used. Before and after treatment, pittsburgh sleep quality index (PSQI), whole night polysomnography, sleep latency (SL), awakening times (AT), sleep efficiency (SE), rapid eye movement (REM), latency (RL), total actual sleep time (TST) and proportion of N1, N2, N3 and REM in the whole sleep stage were recorded. Scores of self-rating depression scale (SDS), self-rating anxiety scale (SAS), kidney Yin deficiency syndrome score and treatment emergent symptom scale (TESS) were graded, and levels of 5-hydroxytryptamine (5-HT), 5-hydroxyindole acetic acid(5-HIAA), gamma-aminobutyric acid (GABA), norepinephrine (NE) and dopamine (DA) were detected. Result:In rank sum test, clinical efficacy in observation group was better than that in control group (Z=2.115, PPPPPPPPχ2=9.945, PConclusion:Addition and subtraction therapy of Liuwei Dihuangwan combined with Huanglian Ejiaotang can improve sleep quality, prolong sleep time, alleviate depression, anxiety, and can also regulate neurotransmitters to improve sleep effect. The efficacy of PSQI is better than that of Estazolam tablets.
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Objective:To study the effect of Liuwei Dihuangwan on the improvement of autophagy level of hippocampal neurons in mice with kidney deficiency Alzheimer' s disease (AD) and its partial mechanism, in order to explore part of therapeutic mechanisms of kidney-tonifying and essence-filling therapy for AD. Method:Healthy male C57-B6 mice were divided into control group, AD group, kidney deficiency AD group and Liuwei Dihuangwan group(1.08 g·kg-1). The control group and the AD group were subcutaneously injected with normal saline (15 mL·kg-1) daily, and the kidney deficiency AD group and the Liuwei Dihuangwan group were subcutaneously injected with hydrocortisone injection (15 mL·kg-1) daily for 20 consecutive days. On the 21st day, the other three groups were injected with 6 μg amyloid beta protein 25-35(Aβ25-35) in the lateral ventricle, while the control group was injected with sterile saline into the lateral ventricle. The levels of serum cortisol and testosterone in each group were detected by enzyme-linked immunosorbent assay (ELISA), the morphological changes in hippocampal neurons were observed by transmission electron microscopy, the expression of microtubule-associated protein 1 light chain 3 (LC3) was detected by immunofluorescence, and the expression of selective autophagic junction protein (p62) was detected by Western blot. Result:Compared with normal group, serum cortisol and testosterone levels in AD group and kidney deficiency AD group were significantly reduced (PPPPPPPPPConclusion:Kidney-tonifying and essence-filling therapy can protect hippocampal neurons, increase LC3 expression in hippocampal neurons, decrease p62 expression level and increase autophagy level of hippocampal neurons. It has a certain therapeutic effect on kidney-deficiency Alzheimer' s disease.
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Objective: To investigate the effect of Liuwei Dihuangwan on connexin 32 (Cx32) in hepatoma cell line CBRH7919 and its gap junction intercellular communication (GJIC), and furthermore study its mechanism of enhancing the bystander killing effect of suicide gene therapy. Method: Liuwei Dihuangwan (32 g·kg·d-1) and the same volume of normal saline were given to the rats by intragastrical administration. Blood was taken to prepare the medicated serum of Liuwei Dihuangwan and blank control serum, respectively. The hepatoma cell line CBRH7919 were treated by control serum and medicated serum of Liuwei Dihuangwan in different concentrations. There were four groups in experiment:the blank control group (volume fraction of 10%), medicated serum high dose group of Liuwei Dihuangwan (the volume fraction of 10%), medicated serum middle dose group of Liuwei Dihuangwan (the volume fraction of 5%), and medicated serum low dose group of Liuwei Dihuangwan (the volume fraction of 2.5%). The expression levels of Cx32 protein and mRNA in hepatoma cell line CBRH7919 were detected by indirect immunofluorescence assay (ⅡA) and Real-time fluorescence quantitative polymerase chain reaction (Real-time PCR) assay. The fluorescence redistribution after photobleaching (FRAP) method was used to detect the function of GJIC of hepatoma cell line CBRH7919. Result: ① The indirect immunofluorescence assay (ⅡA) analysis indicated that as compared with the blank control group, the cx32 expression of CBRH7919 cells was up-regulated in a concentration-dependent manner in each dose group of the serum containing Liuwei Dihuangwan (PPPPConclusion: The mechanism of medicated serum of Liuwei Dihuangwan in enhancing the bystander killing effect of suicide geneis related to gap junction. Liuwei Dihuangwan may enhance the function of GJIC by increasing the localization of cx32 on the cell membrane of CBRH7919 cells and increasing the expression of cx32 mRNA and protein to achieve the synergistic action.